Dimer Organization of Membrane‐Associated NS5A of Hepatitis C Virus as Determined by Highly Sensitive 1H‐Detected Solid‐State NMR,Angewandte Chemie International Edition

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Dimer Organization of Membrane‐Associated NS5A of Hepatitis C Virus as Determined by Highly Sensitive 1H‐Detected Solid‐State NMR
Angewandte Chemie International Edition ( IF 16.1 ) Pub Date : 2020-11-18 , DOI: 10.1002/anie.202013296
Vlastimil Jirasko ₁ , Alons Lends ₁ , Nils-Alexander Lakomek ₁ , Marie-Laure Fogeron ₂ , Marco E Weber ₁ , Alexander A Malär ₁ , Susanne Penzel ₁ , Ralf Bartenschlager _{3,

4} , Beat H Meier ₁ , Anja Böckmann ₂

Affiliation

The Hepatitis C virus nonstructural protein 5A (NS5A) is a membrane‐associated protein involved in multiple steps of the viral life cycle. Direct‐acting antivirals (DAAs) targeting NS5A are a cornerstone of antiviral therapy, but the mode‐of‐action of these drugs is poorly understood. This is due to the lack of information on the membrane‐bound NS5A structure. Herein, we present the structural model of an NS5A AH‐linker‐D1 protein reconstituted as proteoliposomes. We use highly sensitive proton‐detected solid‐state NMR methods suitable to study samples generated through synthetic biology approaches. Spectra analyses disclose that both the AH membrane anchor and the linker are highly flexible. Paramagnetic relaxation enhancements (PRE) reveal that the dimer organization in lipids requires a new type of NS5A self‐interaction not reflected in previous crystal structures. In conclusion, we provide the first characterization of NS5A AH‐linker‐D1 in a lipidic environment shedding light onto the mode‐of‐action of clinically used NS5A inhibitors.

中文翻译：

通过高灵敏 1H 检测固态 NMR 测定丙型肝炎病毒膜相关 NS5A 的二聚体组织

丙型肝炎病毒非结构蛋白 5A (NS5A) 是一种膜相关蛋白，参与病毒生命周期的多个步骤。针对 NS5A 的直接作用抗病毒药物 (DAA) 是抗病毒治疗的基石，但人们对这些药物的作用方式知之甚少。这是由于缺乏膜结合 NS5A 结构的信息。在此，我们提出了重组为蛋白脂质体的 NS5A AH-linker-D1 蛋白的结构模型。我们使用高灵敏度的质子检测固态核磁共振方法，适合研究通过合成生物学方法产生的样品。光谱分析表明 AH 膜锚和连接体均具有高度柔性。顺磁弛豫增强 (PRE) 揭示了脂质中的二聚体组织需要一种新型的 NS5A 自相互作用，而这种相互作用在以前的晶体结构中没有反映出来。总之，我们提供了 NS5A AH-linker-D1 在脂质环境中的首次表征，为临床使用的 NS5A 抑制剂的作用模式提供了线索。

更新日期：2020-11-18

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